Hematocrit measurement in an automatic hematology analyzer

ABSTRACT

In a hematology analyzer, peaks of pulses indicative of red blood cells in a sample during a count are detected. Peak detecting means produce a gate signal connected to gate means which couple a portion of each pulse in the time vicinity of its peak to integration means. The level accumulated in the integration means is indicative of total cell volume, and hence indicative of hematocrit for a known sample volume.

This is a division of application Ser. No. 535,495 filed on Dec. 23,1974 and now U.S. Pat. No. 4,052,596.

This invention relates to a hematology analyzer, and more particularly,to an improvement to such an analyzer for providing output informationrelating to the hematocrit, the mean corpuscular volume, the meancorpuscular hemoglobin and the mean corpuscular hemoglobin concentrationof a blood sample, in addition to the already provided hemoglobin andred blood cell count information of that sample of blood.

The prior art includes apparatus offered for sale by Hycel, Inc. ofHouston, Texas, known as the Hycel Counter 300, Model HC 300 whichperforms a red blood cell count, a white blood cell count and ahemoglobin content determination on a sample of blood. The red bloodcell count may be made by preparing a solution of whole blood in abuffered electrolyte dilutant so that the blood is diluted, forinstance, 160,000 times. A precise volume of this solution is thenpassed through a small opening of approximately 100 microns in diameter,and electrical apparatus connected to the opening monitors the impedanceof the solution flowing therein. As a blood cell passes through theopening, the impedance increases causing a pulse to be provided by theelectrical apparatus. The magnitude of this pulse is proportional to thesize of the cell being measured, and the number of pulses equals thenumber of cells in the precise volume.

In the prior art apparatus, white blood cells are measured insubstantially the same manner as described above for the red blood cellsexcept that a chemical lycing agent is added to distroy the red bloodcells and a larger volume of solution is measured, or on the other hand,a less diluted solution is measured. It should be noted that in the casewhere the red blood cells are being measured, approximately one or twopercent of those detected cells will be white blood cells. Furthermore,the prior art apparatus measures the hemoglobin content of the blood bymixing a specific chemical with the blood and thereafter performing acolormetric determination in a known manner. The output of the photocellused in the colormetric determination is analog voltage in which themagnitude is proportional to the hemoglobin content.

In addition to the red blood cell count, the white blood cell count andthe hemoglobin content value of the blood, a physician may desire toknow the hematocrit value of the blood, the mean corpuscular volume ofthe blood, the mean corpuscular hemoglobin of the blood and the meancorpuscular hemoglobin concentration of the blood.

The hematocrit of the blood is the percent of the blood cell volume tothe total amount of blood of the sample. The classical manner ofdetermining the hematocrit is to add an anticoagulant to the bloodsample and centrifuge it until the cells and serum are separated withthe cells settling to the bottom and the serum on the top. Then a volumemeasurement of the cells and the serum is made and the percentage of thecells to the total is determined as a hematocrit value. A typical valueof the mean corpuscular volume may be between 84 and 95 cubic microns.

The mean corpuscular hemoglobin is the amount of hemoglobin per redblood cell of the blood and may be determined by dividing ten times thehemoglobin in grams per liter by the number of red blood cells per cubicmillimeter of blood and may be expressed in units of micro-micrograms. Atypical value for the mean corpuscular hemoglobin would be 28 to 32micro-micrograms.

The mean corpuscular hemoglobin concentration is the amount ofhemoglobin expressed as a percent of the volume of a red blood cell andmay be calculated by dividing the hemoglobin by the hematocrit. Theresult is expressed in terms of percentage, with a typical value beingin the range of 33-38%.

In order for the hematology analyzer to provide all the desiredinformation to the physician, it should provide a red blood cell count,a white blood cell count, a hemoglobin value, a hematocrit value, a meancorpuscular volume value, a mean corpuscular hemoglobin value, and amean corpuscular hemoglobin concentration value. The prior art deviceprovides only the first three of these values. The red and white bloodcell counts are made by applying the pulses provided when monitoring theimpedance of the flow cell to a digital counter. After a precise volumepasses through the flow cell, the count of the counter is provided to adigital display where it may be read by the operator. In the case of thehemoglobin, the analog voltage provided as a result of the hemoglobincolormetric determination is scaled and applied to the data input of ananalog to digital converter and a direct current reference voltage isconnected to the reference input. The digital output, is then providedto the digital display upon command of the operator.

In order to utilize the prior art device to provide the additional fourvalues, it is necessary to determine the hematocrit value and providedividing means to be able to divide the hematocrit and hemoglobin by thenumber of red blood cells and to divide the hemoglobin by thehematocrit.

U.S. Pat. No. 3,439,267, entitled "Automatic Hematocrit MeasuringApparatus" discloses apparatus for determining the hematocrit value in adiluted solution of whole blood, and the U.S. Pat. No. 3,473,010entitled "Apparatus and Method for Determining Mean Particle Volume"discloses apparatus for determining the mean corpuscular volume of theblood. The apparatus disclosed by both of these patents must be usedtotally in addition to the prior art analyzer described above. It wouldbe more desirable to provide apparatus which can use at least someexisting part of the prior art apparatus, where possible, in order toreduce the cost involved in providing the additional four values.

In accordance with one preferred embodiment of this invention, there isprovided a circuit for selectively providing a digital signalmanifesting a ratio of selected ones of a plurality of analog signalscomprising an analog-to-digital converter means having a data input, areference input and a digital output. The digital output provides adigital signal manifesting a value relating to the analog signal appliedto the data input divided by the analog signal applied to the referenceinput. In addition, there is provided switching means for connecting theselected analog signals to the data and reference inputs of theconversion means to provide the selected digital signals.

One preferred embodiment of this invention is described hereinafter withreference being made to the following FIGURES. in which:

FIG. 1 shows a block diagram of the hematology analyzer for measuringseven hematological parameters of blood;

FIG. 2 shows a series of waveform signals useful in understanding theoperation of the analyzer shown in FIG. 1; and

FIG. 3 shows a circuit diagram of the integrator shown in FIG. 1.

Referring now to FIG. 1, hematology analyzer 10 includes red blood celldetector 12 which provides a pulse signal to amplifier 14. Detector 12may be a flow cell type apparatus, known in the prior art, in which aprecise volume of diluted blood is allowed to flow through a smallopening of, for instance, 100 microns in diameter and the impedance ofthat opening is constantly monitored. Each time a red blood cell entersthe opening, the impedance changes, causing the output voltage appliedthrough the blood in the opening to change amplitude. The amount ofamplitude change depends upon the impedance in the flow cell and thatimpedance, in turn, depends upon the size of the red blood cell.Accordingly each time a blood cell is in the opening, a pulse signal isprovided on the electrical line monitoring the impedance change. Theduration of each pulse depends upon the time a blood cell is in the flowcell and this duration is variable, although typically about thirtymicro-seconds.

After the voltage from detector 12 is amplified by amplifier 14, itappears as waveform A shown in FIG. 2. It should be noted that there isno periodic time cycle of occurrence of the pulses and they havediffering amplitudes reflecting the different size cells.

Comparator 16 responds to waveform A by providing a pulse signal, whichis substantially square shaped, as indicated by wave form B, during thetime the amplitude of the waveform A exceeds a threshold voltage ^(V) T.The output from comparator 16 is provided to a decade counter 18 whichcounts the number of pulses applied thereto during the time a controlsignal so permits. The control signal may be provided from means (notshown) during the time that the volume of blood having cells counted ispassed through the flow cell opening. After the control signal indicatesthe counting is complete, the count in decade counter 18 indicates thenumber of red blood cells counted for the precise volume of blood beingmeasured.

Analyzer 10 also includes a hemoglobin detector 20 which provides ananalog voltage HGB manifesting the value of the hemoglobin of the sampleof blood being tested. This voltage is provided by a photocell used inmaking the colormetric measurement at a certain wavelength of a solutionof the blood and other chemicals in a manner well known in the art.Detector 20 may also include amplification and scaling means forproviding the HGB signal at a proper magnitude.

The B signal from comparator 16 is also applied to a binary counter 22which counts the number of B pulses during the time the control signalapplied to counter 22 so allows. The binary count in counter 22 isapplied to a digital-to-analog converter 24, which converts the digitalcount manifested at the output of counter 22 into an analog voltage RBCmanifesting the number of red blood cells detected and counted.

To determine the hematocrit value, the A signal at the output ofamplifier 14 is provided to a differentiating circuit 26, which providesthe signal shown as waveform C in FIG. 2. Waveform C has, for each cellpulse, a leading positive pulse and trailing negative pulse and a zerocrossing between the two pulses which occurs at the time of maximumamplitude of the A signal pulses. The output from the differentiatingcircuit 26 is applied to a zero cross detector circuit 28 which provideswaveform D shown in FIG. 2. Waveform D is a narrow trigger pulse signalapplied each time the amplitude of the differentiated C signal crosseszero. The output from zero cross detector circuit 28 is applied to theset input to trigger monostable multivibrator 30 if the B pulse appliedto the inhibit input thereof is occurring. Multivibrator 30 provides theE signal shown in FIG. 2, that is, a fixed duration pulse of, forinstance, three microseconds. The E signal is applied to the gate offield effect transistor 32 to render it conductive.

The A signal from amplifier 14 is connected to one main electrode offield effect transistor 32, the other main electrode of which isconnected to integrator 34. When multivibrator 30 is triggered, therebyrendering transistor 32 conductive, waveform F is provided to integratorcircuit 34.

Waveform F has approximately straight line sides, an upper portion equalto the magnitude of waveform A and a duration equal to the duration ofthe monostable multivibrator 30 output pulse. Integrator circuit 34integrates each of the F pulses applied thereto, and provides an analogvoltage signal HCT, shown as waveform G in FIG. 2, which is equal to theintegral of the F waveform. This voltage is proportional to thehematocrit value.

Referring now to FIG. 3, a circuit diagram of integrator 34 is shown andincludes an operational amplifier 36 having an input 38 and an output40. Connected between input 38 and output 40 is a capacitor 42 andconnected between transistor 32 and input 38 is a variable resistor 44.The value of capacitor 42 and the value set for resistor 44 scales thevoltage provided by integrator 34 and these values are selected to causethe voltage at output 40 to manifest the hematocrit value. In useresistor 44 may be adjusted by the operator during a calibrating mode ofoperation when a sample having a known hematocrit value is used.

In addition, analyzer 10 includes an analog-to-digital converter 46having a data input and reference input. Analog signals are provided toboth the data input and the reference input and a digital output signalappears at the output indicated schematically by five output digitallines. The digital value manifested by the output signal from converter46 is equal to the analog signal applied to the data input divided bythe analog signal applied to the reference input.

To couple the proper signals to the analog-to-digital converter 46, adouble ganged rotary switch 48 is provided. Rotary switch 48 has twooutput terminals 50 and 52 and two sets of five input terminalscorrespondingly associated with each output terminal and respectivelydesignated as HGB, HCT, MCHC, MCH, and MCV. A switching arm connected toeach output terminal selectively connects the same corresponding one ofthe two sets of input terminals to that output terminal so that thefollowing respectively ordered determinations are made: Hemoglobin,hematocrit, mean corpuscular hemoglobin, mean corpuscular hemoglobinconcentration and mean corpuscular volume. The HGB signal is connectedto the HGB, MCHC and MCH input terminals associated with output terminal50. The RBC signal is connected to the MCH and MCV input terminalsassociated with output terminal 52. The HCT signal is connected to theHCT and MCV input teminals associated with the output terminal 50 and tothe MCHC input terminal associated with output terminal 52. A referencedirect current voltage of, for instance, 5 volts, is connected to theHGB and HCT input terminals associated with output terminal 52.

Connected in this manner, when the two switching arms connect the HGBinput terminals to the output terminals 50 and 52, the value manifestedat the output of analog-to-digital converter 46 is the hemoglobin value.When the switching arms connect the HCT input terminals to the outputterminals 50 and 52, the value manifested at the output of converter 46is the hematocrit value. When the switching arms connect the MCHC inputterminals to the output terminals 50 and 52, the value manifested at theoutput of converter 46 is the mean corpuscular hemoglobin concentrationvalue. When the switching arms connect the MCH input terminals to theoutput terminals 50 and 52, the signal manifested at the output ofconverter 46 is the mean corpuscular hemoglobin. Finally, when theswitching arms connect the MCV input terminals to the output terminals50 and 52, the signal manifested at the output of converter 46 is themean corpuscular volume.

The digital outputs from converter 46 and counter 18 are applied to adigital display 54 which provides a numerical display manifesting thedigital numbers applied thereto. Display 54 may be under the control ofsignals (not shown) causing it to display the desired signals uponcommand of the operator.

What is claimed is:
 1. In a blood cell analyzer in which a cell signalis provided in response to the detection of blood cells in a precisevolume of blood, said cell signal including a plurality of pulses, witheach pulse representing the detection of one cell and with the magnitudeof each pulse relating to the size of each detected cells, hematocritmeasuring means for providing a voltage relating to the hematocrit ofthe blood comprising:integration means for summing signals appliedthereto; controlled switching means for coupling said pulses tointegration means; and detecting means for detecting the maximum ofmagnitude of each pulse of said cell signal and for producing a gatesignal indicative of said detected maximum magnitude and having a fixed,preselected width which is narrow compared to a typical pulse width,said detecting means being coupled to said controlled switching meansfor rendering said controlled switching means conductive in response tosaid gate signal, whereby said integrator is incremented by a voltageindicative of a peak portion of each pulse and the voltage stored insaid integration means at the completion of said cell signal comprisesthe voltage relating to the hematocrit of the blood.
 2. The hematocritmeasuring means according to claim 1 wherein said detecting meanscomprises a differentiating circuit providing a differential signal andmeans connected to said differentiating circuit for producing said gatesignal when said differential signal reaches a preselected value.
 3. Thehematocrit measuring means according to claim 2 wherein said means forproducing said gate signal comprises a zero crossing detector and amonostable multivibrator connected for triggering by a trigger signalfrom said zero crossing detector, the output of said multivibratorcomprising said gate signal.
 4. The hematocrit measuring means accordingto claim 2 further comprising means for inhibiting said multivibratorwhen said pulse is below a preselected threshold level.
 5. Thehematrocrit measuring means according to claim 4 further comprisingmeans for comparing the voltage stored in said integration means at thecompletion of said cell signal to a reference voltage and converting theresult of said comparison to a digital hematocrit indication.